Chromatin immunoprecipitation was performed with 35 μg of cross-linked chromatin from HeLa cells treated by TNF-α (30ng/ml, 1h) using 5 μg of STAT1 Rabbit mAb (A19563). DNA libraries were prepared using Scale ssDNA-seq Lib Prep Kit for Illumina V2 (RK20228). The ChIP sequencing results indicate the enrichment pattern of STAT1 across chromosome 6 (upper panel) and the genomic region encompassing TAP1, a representative gene enriched in STAT1 (lower panel).
Chromatin immunoprecipitation was performed with 35 μg of cross-linked chromatin from HeLa cells treated by TNF-α (30ng/ml, 1h) using 5 μg of STAT1 Rabbit mAb (A19563). DNA libraries were prepared using Scale ssDNA-seq Lib Prep Kit for Illumina V2 (RK20228). The ChIP sequencing results indicate the enrichment pattern of STAT1 in the representative genomic region surrounding TAP1 gene.
Western blot analysis of various lysates using STAT1 Rabbit mAb (A19563) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 1s.
Immunoprecipitation analysis of 600 μg extracts of HeLa cells using 3 μg STAT1 antibody (A19563). Western blot was performed from the immunoprecipitate using STAT1 antibody (A19563) at a dilution of 1:1000.
Western blot analysis of extracts of Mouse lung cells, using JAK1 antibody (A11963) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Enhanced Kit (RM00021).
Exposure time: 180s.
Immunohistochemistry analysis of paraffin-embedded rat ovary using JAK1 Rabbit mAb (A11963) at dilution of 1:100 (40x lens).Perform microwave antigen retrieval with 10 mM PBS buffer pH 7.2 before commencing with IHC staining protocol.
Immunohistochemistry analysis of paraffin-embedded human colon carcinoma using JAK1 Rabbit mAb (A11963) at dilution of 1:100 (40x lens).Perform microwave antigen retrieval with 10 mM PBS buffer pH 7.2 before commencing with IHC staining protocol.
Immunohistochemistry analysis of paraffin-embedded mouse spleen using JAK1 Rabbit mAb (A11963) at dilution of 1:100 (40x lens).Perform microwave antigen retrieval with 10 mM PBS buffer pH 7.2 before commencing with IHC staining protocol.
Western blot analysis of extracts of Raji cells, using SHP1 antibody (A19111) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 10s.
Western blot analysis of extracts of various cell lines, using SHP1 antibody (A19111) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 3min.
Immunohistochemistry analysis of SHP1 in paraffin-embedded human spleen tissue using SHP1 Rabbit mAb (A19111) at a dilution of 1:200 (40x lens).High pressure antigen retrieval was performed with 0.01 M citrate buffer (pH 6.0) prior to IHC staining.
Immunohistochemistry analysis of SHP1 in paraffin-embedded mouse spleen tissue using SHP1 Rabbit mAb (A19111) at a dilution of 1:800 (40x lens).High pressure antigen retrieval was performed with 0.01 M citrate buffer (pH 6.0) prior to IHC staining.
Immunohistochemistry analysis of SHP1 in paraffin-embedded human tonsil tissue using SHP1 Rabbit mAb (A19111) at a dilution of 1:200 (40x lens).High pressure antigen retrieval was performed with 0.01 M citrate buffer (pH 6.0) prior to IHC staining.
Immunohistochemistry analysis of SHP1 in paraffin-embedded rat spleen tissue using SHP1 Rabbit mAb (A19111) at a dilution of 1:800 (40x lens).High pressure antigen retrieval was performed with 0.01 M citrate buffer (pH 6.0) prior to IHC staining.
Immunofluorescence analysis of rat spleen using SHP1 Rabbit mAb (A19111) at dilution of 1:100 (40x lens). Blue: DAPI for nuclear staining.
Immunofluorescence analysis of mouse spleen using SHP1 Rabbit mAb (A19111) at dilution of 1:100 (40x lens). Blue: DAPI for nuclear staining.
Western blot analysis of various lysates using Phospho-STAT1-Y701 Rabbit mAb (AP0054) at 1:1000 dilution.HeLa,NIH/3T3 and PC-12 cells were treated by IFN-α (100 ng/ml) at 37℃ for 30 minutes after serum-starvation overnight.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 180s.
Western blot analysis of lysates from K-562 cells, using SOCS2 Rabbit mAb (A9190) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 1s.
Western blot analysis of various lysates using SOCS2 Rabbit mAb (A9190) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Enhanced Kit (RM00021).
Exposure time: 3min.
Immunohistochemistry analysis of SOCS2 in paraffin-embedded rat lung using SOCS2 Rabbit mAb (A9190) at dilution of 1:100 (40x lens).Perform microwave antigen retrieval with 10 mM Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Confocal imaging of C2C12 cells using SOCS2 Rabbit mAb (A9190, dilution 1:200) followed by a further incubation with Cy3 Goat Anti-Rabbit IgG (H+L) (AS007, dilution 1:500)(Red). The cells were counterstained with α-Tubulin Mouse mAb (AC012, dilution 1:400) followed by incubation with ABflo® 488-conjugated Goat Anti-Mouse IgG (H+L) Ab (AS076, dilution 1:500) (Green). DAPI was used for nuclear staining (Blue). Objective: 100x.
Confocal imaging of MCF7 cells using SOCS2 Rabbit mAb (A9190, dilution 1:200) followed by a further incubation with Cy3 Goat Anti-Rabbit IgG (H+L) (AS007, dilution 1:500) (Red). The cells were counterstained with α-Tubulin Mouse mAb (AC012, dilution 1:400) followed by incubation with ABflo® 488-conjugated Goat Anti-Mouse IgG (H+L) Ab (AS076, dilution 1:500) (Green). DAPI was used for nuclear staining (Blue). Objective: 100x.
Western blot analysis of various lysates using Phospho-STAT3-S727 Rabbit mAb (AP0715) at 1:1000 dilution.NIH/3T3 and PC-12 cells were treated by TNF-α (20 ng/mL) at 37℃ for 30 minutes.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 120s.
Immunohistochemistry analysis of Phospho-STAT3-S727 in paraffin-embedded mouse kidney using Phospho-STAT3-S727 Rabbit mAb (AP0715) at dilution of 1:200 (40x lens).Perform high pressure antigen retrieval with 10 mM citrate buffer pH 6.0 before commencing with IHC staining protocol.
Immunohistochemistry analysis of Phospho-STAT3-S727 in paraffin-embedded mouse liver using Phospho-STAT3-S727 Rabbit mAb (AP0715) at dilution of 1:200 (40x lens).Perform high pressure antigen retrieval with 10 mM citrate buffer pH 6.0 before commencing with IHC staining protocol.
Immunohistochemistry analysis of Phospho-STAT3-S727 in paraffin-embedded rat colon using Phospho-STAT3-S727 Rabbit mAb (AP0715) at dilution of 1:200 (40x lens).Perform high pressure antigen retrieval with 10 mM citrate buffer pH 6.0 before commencing with IHC staining protocol.
Immunohistochemistry analysis of Phospho-STAT3-S727 in paraffin-embedded rat liver using Phospho-STAT3-S727 Rabbit mAb (AP0715) at dilution of 1:200 (40x lens).Perform high pressure antigen retrieval with 10 mM citrate buffer pH 6.0 before commencing with IHC staining protocol.
Western blot analysis of various lysates using STAT4 Rabbit mAb (A4523) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 90s.
Immunofluorescence analysis of C6 cells using STAT4 Rabbit mAb (A4523) at dilution of 1:100 (40x lens). Secondary antibody: Cy3 Goat Anti-Rabbit IgG (H+L) (AS007) at 1:500 dilution. Blue: DAPI for nuclear staining.
Immunofluorescence analysis of NIH-3T3 cells using STAT4 Rabbit mAb (A4523) at dilution of 1:100 (40x lens). Secondary antibody: Cy3 Goat Anti-Rabbit IgG (H+L) (AS007) at 1:500 dilution. Blue: DAPI for nuclear staining.
Immunoprecipitation analysis of 300 μg extracts of HeLa cells using 3 μg STAT4 antibody (A4523). Western blot was performed from the immunoprecipitate using STAT4 antibody (A4523) at a dilution of 1:1000.
Western blot analysis of extracts of various cell lines, using STAT6 antibody (A19120) at 1:1000 dilution.
Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) (AS014) at 1:10000 dilution.
Lysates/proteins: 25μg per lane.
Blocking buffer: 3% nonfat dry milk in TBST.
Detection: ECL Basic Kit (RM00020).
Exposure time: 3min.
Immunofluorescence analysis of HeLa using STAT6 Rabbit mAb (A19120) at dilution of 1:100 (40x lens). Blue: DAPI for nuclear staining.
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